The anion exchange protein (band 3) is an abundant protein in the human[unreadable] erythrocyte membrane which is composed of two domains that serve[unreadable] distinct functions. The integral membrane domain facilitates the[unreadable] exchange of bicarbonate and chloride, thereby increasing the CO2[unreadable] carrying capacity of the blood. The cytoplasmic domain serves as a[unreadable] binding site for peripheral proteins including ankyrin and band 4.2[unreadable] which are involved in forming a physical link between the plasma[unreadable] membrane and the membrane skeleton. These protein-protein interactions[unreadable] are essential for maintaining the unique biconcave shape and[unreadable] viscoelastic properties of erythrocytes. Genetic defects in band 3,[unreadable] ankyrin, or band 4.2 result in abnormal cell shapes, rigid or fragile[unreadable] cells, and often, clinically significant hemolytic anemias. The long-[unreadable] term goals of these studies are to determine the structural and dynamic[unreadable] properties of band 3 that are essential for its role as an organizing[unreadable] center for protein-protein interactions which stabilize the erythrocyte[unreadable] membrane.[unreadable] [unreadable] It is hypothesized that band 3 is present primarily as dimers and[unreadable] tetramers, that these small oligomers interact with the membrane[unreadable] skeleton via a flexible linkage, and that the flexibility of the[unreadable] cytoplasmic domain of band 3 is necessary for normal cell shape and[unreadable] mechanical properties. In the next grant period, electron paramagnetic[unreadable] resonance, fluorescence, and phosphorescence spectroscopies will be[unreadable] combined with classical biochemical methods to address the following[unreadable] specific questions: 1) What are the sizes of the oligomeric species of[unreadable] band 3 present in the erythrocyte membrane? 2) How flexible, in terms[unreadable] of the amplitude of motion, is the segment of the cytoplasmic domain of[unreadable] band 3 which links the membrane skeleton to the lipid bilayer, and to[unreadable] what extent do these links exist in the intact cell? 3) What are the[unreadable] properties of specific protein-protein interactions involving the[unreadable] cytoplasmic domain of band 3 and the binding proteins ankyrin and band[unreadable] 4.2? To help accomplish these goals, new molecular probes,[unreadable] spectroscopic methods, and data analysis methods will be developed which[unreadable] will be of utility in the biomedical research community.[unreadable]